COMPARATIVE-STUDY OF DISTRIBUTION AND BIOCHEMICAL-CHARACTERIZATION OF BRAIN CALCITONIN GENE-RELATED PEPTIDE RECEPTORS IN 5 DIFFERENT SPECIES

Calcitonin gene-related peptide receptors are abundant in the brain, and the pattern of distribution of receptors is similar in the rat, pig, cow, sheep and in man. In comparison to the cerebellum (100%), only a small number of receptors were found in the ventral spinal cord (20%), cerebral white matter (20%), pituitary (27%) and hypothalamus (33%). In contrast, cerebral cortex (70%), thalamus (50%) and dorsal spinal cord (50%) contained a higher number of calcitonin gene-related peptide receptors. Overall, the highest number of receptors was observed in the pig brain followed by the rat, human, sheep and cow. IC50 values of alpha- and beta-calcitonin gene-related peptide for cerebellar membranes were 200 pM, while its antagonist calcitonin gene-related peptide(8-37) and (9-37) had an IC50 is similar to 1 nM. The recently discovered 37-amino acid peptide amylin (46% homology with calcitonin gene-related peptide) displaces the membrane bound [I-125]calcitonin gene-related peptide with 50-fold molar excess (IC50 = 10 nM). The only other peptide able to compete for calcitonin gene-related peptide receptor binding is salmon calcitonin, at a > 1000-fold molar excess (IC50 = 250 nM). Dissociation of [I-125]calcitonin gene-related peptide from cerebellar membranes was biphasic, suggesting that calcitonin gene-related peptide receptors in the brain were heterogeneous. Scatchard analysis of the binding data revealed significant differences in binding capacities (B(max)) and affinities (K(d)) of calcitonin gene-related peptide receptors in the cerebellum of these species. Scatchard plots obtained from [I-125]calcitonin gene-related peptide binding to brain membranes from all five species were curvilinear, and suggested the presence of both high- and low-affinity binding sites. Among the high-affinity binding sites for [I-125]calcitonin gene-related peptide, the highest was found in the pig cerebellum (K(d) = 0.2 nM, B(max) = 12 fmol/mg) and the lowest in the cow (K(d) = 3.4 nM, B(max) = 8.2 fmol/mg). The K(d) of the low-affinity binding sites varied from 5.1 to 21.2 nM and B(max) from 110 to 225 fmol/mg protein. The highest number of calcitonin gene-related peptide specific bindings sites (receptors) was found in the cerebellum of the pig, followed by the rat, man, sheep and cow. The large numbers of calcitonin gene-related peptide receptors found in the cerebral blood vessels of all species suggests a role of calcitonin gene-related peptide in the regulation of cerebral blood flow. This study demonstrates the high similarity of the distribution and some of the biochemical characteristics of calcitonin gene-related peptide receptors in five species.